Second-generation octarellins: two new de novo (beta/alpha)8 polypeptides designed for investigating the influence of beta-residue packing on the alpha/beta-barrel structure stability
[en] The sequence of octarellin I, the first de novo (beta/alpha)8 polypeptide, was revised according to several criteria, among others the symmetry of the sequence, beta-residue volume and hydrophobicity, and charge distribution. These considerations and the overall conclusions drawn from the first design led to two new sequences, corresponding to octarellins II and III. Octarellin II retains perfect 8-fold symmetry. Octarellin III has the same sequence as octarellin II, except for the beta-strands which exhibit a 4-fold symmetry. The two proteins were produced in Escherichia coli. Infrared and CD spectral analyses of octarellins II and III reveal a high secondary structure content. Non-denaturing gel electrophoresis, molecular sieve chromatography and analytical ultracentrifugation suggest that both of these second-generation artificial polypeptides exist as a mixture of a monomer and a dimer form. Octarellins II and III are at least 10 times more soluble than octarellin I. Urea-induced unfolding followed by fluorescence emission suggests that the tryptophan residues, designed to be buried in the (beta/alpha)8, are indeed packed in the hydrophobic core of both proteins. However, octarellin III displays a higher stability towards urea denaturation, indicating that introducing 4-fold symmetry into the beta-barrel might be important for stability of the overall folding.
Mainfroid, Véronique; Université de Liège - ULiège
Preaux, Gisèle; Katholieke Universiteit Leuven - KUL
Lamproye, Alain; Eurogentec
Poncin, Alain; Eurogentec
Goormaghtigh, Erik; Université Libre de Bruxelles - ULB
Ruysschaert, Jean-Marie; Université Libre de Bruxelles - ULB
Martial, Joseph ; Université de Liège - ULiège > Département des sciences de la vie > GIGA-R : Biologie et génétique moléculaire
Goraj, Karine; Université de Liège - ULiège
Language :
English
Title :
Second-generation octarellins: two new de novo (beta/alpha)8 polypeptides designed for investigating the influence of beta-residue packing on the alpha/beta-barrel structure stability
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